1. Field of the Invention
Embodiments of the present invention concern antibodies binding MUC18 antigen as well as methods and means for making and using such antibodies.
2. Description of the Related Art
MUC18 is a cell-surface glycoprotein originally identified as a melanoma antigen, melanoma cell adhesion molecule (MCAM), whose expression is associated with tumor progression and the development of metastatic potential. MUC18 is a 113 kDA cell surface integral membrane glycoprotein composed of a signal peptide, five immunoglobulin-like domains, a transmembrane region, and a short cytoplasmic tail (Lehmann et al., Proc Natl Acad Sci USA, 86(24):9891–5 (1989)).
MUC18 is a member of the immunoglobulin superfamily and has significant sequence homology to a number of cell adhesion molecules of the Ig superfamily (Lehmann et al., Proc. Natl. Acad. Sci. USA, 86:9891–9895 (1989)), including BEN (Pourquie et al., Proc. Natl. Acad. Sci. USA, 89:5261–5265 (1992)), neural-cell adhesion molecule (N-CAM) (Owens et al., Proc. Natl., Acad. Sci. USA, 84:294–298 (1987)), myelin-associated glycoprotein (MAG) (Lai et al., Proc. Natl. Acad. Sci. USA, 84:4337–4341 (1987)), deleted in colorectal cancer (DCC) (Hedrick et al., Genes Devel., 8(10):1174–83 (1994)), and gicerin (Taira et al., Neuron, 12: 861–872 (1994)). The expression of MUC18 has been detected in relatively limited spectrum of normal human tissues and in a variety of malignant neoplasms. In normal adult tissues, MUC18 is expressed on endothelial cells, smooth muscle cells (Shih et al., Lab. Invest., 75:377–388 (1996);Sers et al., Cancer Res., 54(21):5689–94 (1994)), a subpopulation of activated T lymphocytes (Pickl et al., J. Immunol., 158:2107–2115 (1997)) and intermediate trophoblasts (Shih et al., Lab. Invest., 75:377–388 (1996)). MUC18 is also expressed on a variety of malignant neoplasms including smooth muscle neoplasms (Leiomyomas and leiomyosarcomas), tumors of vascular origin (angiosarcomas and Kaposi's sarcomas), placental site trophoblastic tumors, choriocarcinomas and melanomas (Shih et al., Clinical Cancer Res., 2:569–575 (1996); Holzmann et al., Int. J. Cancer, 39:466–471 (1987)). The expression of MUC18 correlates directly with the metastatic potential of human melanoma cells (Bar-Eli, M., Cancer Metastasis, 18(3):377–85 (1999)).
A number of studies have identified MUC18 as a marker of tumor progression and metastasis in melanomas. The expression of MUC18 is absent in normal melanocytes and benign nevi but prominent on many primary melanomas and in most metastatic lesions (Lehmann et al., Proc. Natl. Acad. Sci. USA, 86:9891–9895 (1989); Lehmann et al., Cancer Res., 47:841–845 (1987); Shih et al., Cancer Res., 54:2514–2520 (1994)). Importantly, MUC18 expression correlates well with tumor vertical thickness and metastasis formation, and greater than 80% of metastatic lesions express MUC18 (Lehmann et al., Proc. Natl. Acad. Sci. USA, 86:9891–9895 (1989); Xie et al., Cancer Res., 57:2295–2303 (1997); Sers et al., Proc. Natl. Acad. Sci. USA, 90:8514–8518 (1993); Lehmann et al., Cancer Res., 47:841–845 (1987); Shih et al., Cancer Res., 54:2514–2520 (1994). A diagram depicting the expression of MUC18 with respect to other known molecular lesions in human melanoma is presented in FIG. 1.
The expression of the transcription factors ATF-1 and CREB is upregulated in metastatic melanoma cells. However, how overexpression of ATF-1/CREB contributes to the acquisition of the metastasis is unclear. CREB/ATF-1 may play an essential role in invasion by regulating the CRE-dependent expression of the adhesion molecule MUC18 and metalloproteinase MMP-2 (Jean et al., Mol. Cell Biochem., 212(1–2):19–28 (2000)) which belongs to the MMP family known to contribute to cancers and to have a role in tumor invasion, angiogenesis, and metastasis. Tumor cells are believed to utilize the matrix degrading capability of MMPs to spread to distant sites, and once the tumor cells have metastasized, MMPs are thought to promote the growth of these tumor cells. The role of MUC18 in melanoma tumor progression is not completely understood, but may include a role in one or more steps in the metastatic process possibly by affecting MMP-2 activation or cell migration.
The analysis of human melanoma cell lines showed a positive correlation of MUC 18 expression with the ability of cells to produce metastases in nude mice (Johnson et al., Cancer Metastasis Rev., 18:345–357 (1999)). The generation of tumorigenic variants from a non-tumorigenic melanoma cell line was reported to be accompanied by induction of MUC18 expression (Luca et al., Melanoma Res., 3:35–41 (1993)). Expression of MUC18 on MUC18-negative human melanoma cell lines increased their tumorigenicity and enhanced their metastatic capability in experimental tumor models (Xie et al., Cancer Res., 57:2295–2303 (1997); Bani et al., Cancer Res., 56:3075–3086 (1996)). Finally, inhibition of MUC18 expression in metastases using genetic suppressor elements of MUC18 cDNA led to a decrease of the tumorigenic phenotype in nude mice (Styamoorthy et al., Oncogene, 20:4676 (2001)).
Although the function of MUC18 is not fully understood, several studies have demonstrated a role for this protein in mediating cell-cell and cell-matrix interactions by binding to an unidentified ligand (Shih et al., Cancer Res., 57:3835–3840 (1997); Johnson et al., Int. J. cancer, 73:769–774 (1997)). The expression of cell adhesion molecules which mediate cell-to-cell or cell-to-matrix interactions is a tumor cell property that is essential for metastases. Accordingly, MUC18-transfected melanoma cells showed increased homotypic adhesion, increased attachment to human endothelial cells, and increased invasion through Matrigel-coated filters suggesting a role in tumor invasion and trans-endothelial migration (Xie et al., Cancer Res., 57:2295–2303 (1997)). Importantly, anti-MUC 18 antibodies were able to inhibit these functions in the MUC 18-transfected cells (Xie et al., Cancer Res., 57:2295–2303 (1997)). Accordingly, there is a great need for anti-MUC 18 antibodies that are able to inhibit the biological function of MUC18, most importantly cell proliferation and growth which may be essential to tumor progression and metastasis. Such antibodies would likely interfere with the inherent ability of MUC18 to mediate cell-cell and cell-matrix interactions. The inhibition of such activity may be possible with a monoclonal antibody targeted to MUC18. The ability to affect the progression of tumor cells expressing MUC18 on the cell surface may prove to be a treatment for patients with tumors or of use for prevention of metastatic disease in patients with such tumors.